This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. The principle mechanism responsible for the contraction and relaxation of smooth muscle involves the reversible phosphorylation of the 20 kDa regulatory myosin light chain (rMLC). Typically, elevations in intracellular Ca2+ result in rMLC phosphorylation at the serine-19 (or adjacent threonine-18) residue by the Ca2+/calmodulin-activated MLC kinase (MLCK). The studies in this collaboration are intended to extend findings with respect to the physiological importance of this alternate-site phosphorylation in vascular smooth muscle, and the signaling pathways involved. Continued studies are proposed using accurate mass-driven analysis and rapid parallel MS/MS acquisition to further identify and clarify the physiological conditions under which this alternate site rMLC phosphorylation occurs.